DNA Release from lPEI-g-PEG/DNA Nanoparticles

The release of DNA from lPEI-g-PEG/DNA nanoparticles was assessed in the presence of heparin sulfate as modified from our previously reported protocol.⁵³ An aliquot of 20 μL of nanoparticles solution containing 1 μg of DNA was added to each well of a 96-well plate followed by the addition of 80 μL of 1 mg/mL ethidium bromide solution. To this solution, 100 μL of heparin sulfate solution with increasing concentrations in 300 mM NaCl solution was added to each well and mixed thoroughly, giving final heparin sulfate concentrations ranging from 1 μg/mL to 500 μg/mL in 150 mM NaCl. The solutions were incubated at room temperature for 15 min, and the fluorescence intensity (λ_ex = 510 nm, λ_em = 595 nm) was measured using a fluorescence plate reader (SpectraMax Gemini XPS, Molecular Devices, Sunnyvale, CA). The percentage of DNA released was calculated according to a calibration curve of plasmid DNA subjected to the same conditions.